Parkinson's Disease (PD) is a slowly progressive neurodegenerative disease involving the loss of dopamine (DA) neurons in the brain. The prevalence rate of PD is estimated at 0.3% of the population, making it the second most common neurodegenerative disorder following Alzheimer's disease. VA medical centers annually treat 20,000 - 40,000 patients per year at high cost and numbers are predicted to climb as the population ages. In contrast to the modest advances made in symptomatic treatment for PD there is presently no treatment proven to slow or halt the progressive degeneration of DA neurons. In recognition of this need the VA has issued an RFA entitled BLR&D Merit Review Award for Research on Amyotrophic Lateral Sclerosis, Parkinson's Disease and Alzheimer's Disease to study the etiology and pathogenesis of these diseases. Our primary objective in this competing renewal is to identify promising pharmacotherapeutic approaches that can rapidly translate into neuroprotection in PD by building upon our previous success in developing a novel symptomatic treatment for PD. In the previous funding cycle we demonstrated that a serotonin 5-HT1A receptor agonist reduced glutamatergic activity in the basal ganglia. Pre-clinical evidence supports a role for glutamate in exacerbating DA neuron degeneration. It is unclear how glutamate contributes to the development of PD, but one possibility is through nitric oxide production. Another prominent factor in the pathogenesis of PD is the protein, alpha synuclein (ASyn), but again its precise involvement is unclear. We propose that DA neuronal degeneration is caused by an interaction in which excessive glutamatergic or excitotoxic tone modifies ASyn properties and function through nitric oxide. We will test the hypothesis that reducing excitotoxic drive to DA neurons will be neuroprotective by reducing nitric oxide and we will demonstrate that administration of 5-HT1A agonists is one means of achieving this goal. To accomplish this, our lab has developed an in vivo rat model of slowly progressive (occurring over many weeks) DA neuronal degeneration involving targeted upregulation of ASyn. In this model, ASyn upregulation in the substantia nigra pars compacta (SNc) is caused by intranigral microinjection of a recombinant adeno-associated viral vector containing the ASyn gene. This model was initially developed several years ago, but it has not been widely used to study neuroprotective strategies. In Aim I, we will establish that 5-HT1A receptors within basal ganglia circuitry are situated to reduce excitatory drive to the SNc. To accomplish this we will carry out double label immunohistochemistry and retrograde tracing studies. In Aim II we will use our in vivo model to test the neuroprotective efficacy of treatment strategies, including 5-HT1A agonists, aimed at reducing glutamate-mediated activity and glutamate-related mechanisms in the SNc. Animals will be assessed for motor function throughout 18 weeks of observation. At varying time intervals, immunohistochemical and biochemical studies will measure indices of DA neuronal dysfunction, DA cell loss and biochemical modifications of ASyn across treatment groups. In Aim III we will use the in vivo model to demonstrate the importance of nitric oxide in mediating neurodegeneration. Specifically, we will determine how the approaches used in Aim II affect nitric oxide synthase activity and nitrosation of ASyn. For comparison, we will assess the neuroprotective and biochemical effects of two nitric oxide synthase inhibitors as well. These studies will fill a gap in our understanding of the interaction between ASyn and glutamate- related mechanisms that lead to DA neuronal degeneration. This information will also further our understanding of the mechanisms underlying the motor and potential neuroprotective actions of 5-HT1A drugs. Together, the results of these studies will guide us to developing rational strategies for slowing the progressive degeneration of DA neurons in patients suffering from Parkinson's Disease.